1. Field of the Invention
The present invention relates to scanning optical microscopes and, more particularly, to a laser scanning microscope (LSM) that performs focal point movement along the direction of the optical axis by using a wavefront converting element.
2. Discussion of Related Art
It has heretofore been necessary in order to obtain a three-dimensional image of a specimen with an LSM, for example, to capture optical images of successive planes inside the specimen by mechanically moving either the specimen or the objective along the direction of the optical axis. With this method, however, it is difficult to realize positional control with high accuracy and high reproducibility because the method needs mechanical drive. In a case where the specimen is moved, high-speed scanning cannot be effected when the specimen is large in size.
In observation of a biological specimen, if the objective is scanned in the state of being in direct contact with the specimen or immersed in a culture solution of the specimen, vibrations of the objective adversely affect the specimen under observation.
To solve the above-described problems, Japanese Patent Application Unexamined Publication (KOKAI) No. Hei 11-101942 discloses an adaptive optical apparatus. The apparatus is a microscope having an optical element (wavefront converting element) capable of changing power. The arrangement of the microscope is shown in FIGS. 27 and 28. In this prior art, a wavefront converting element is inserted in either or both of a viewing optical path and an illuminating optical path to change the focal length of the optical system and to correct aberration due to the change of the focal length by using the wavefront converting element. With this arrangement, it is possible not only to form and move a focal point in the object space without changing the distance between the objective and the specimen but also to correct aberration.
In the above-described prior art, it is preferable to place the wavefront converting element in the pupil plane of the objective or at a position conjugate to the pupil plane from the viewpoint of allowing the wavefront converting element to effectively perform its functions of moving the focal point in the object space and making aberration correction. If the wavefront converting element is not conjugate to the pupil plane, illuminating light or image-forming light will pass at different positions on the wavefront converting element according to the height of the object detected by the objective. To perform focal point movement or aberration correction, the wavefront shape has to be changed according to the object height. If the wavefront shape cannot properly be changed, image quality is likely to degrade considerably in an area where the object height is high.
If the wavefront converting element is changed into an optimum shape in accordance with a change in the object height, even if the wavefront converting element is not conjugate to the pupil plane, it is possible to avoid image quality degradation in an area where the object height is high. To realize this, however, the wavefront converting element needs to be controlled at high speed so as to provide an optimum rotationally asymmetric configuration. This is extremely difficult.
For the reasons stated above, it is desirable that the wavefront converting element should be placed at a position conjugate to the pupil. This is, however, difficult to implement because of the following problems.
A variety of objectives are used in microscopic observation, and the pupil position differs for each objective. Therefore, when a plurality of objectives are switched from one to another to perform observation, it is difficult to keep the pupils of the objectives in conjugate relation to the wavefront converting element at all times.
Further, the wavefront converting element needs to be placed in conjugate relation to the position of a laser scanning member and also to the position of the objective pupil. Accordingly, at least two pupil relay optical systems are required. Therefore, the apparatus becomes large in size and complicated unfavorably.
Further, in the above-described prior art, a reflection type wavefront converting element is incorporated in the illuminating optical path or/and the light-detecting optical path. Therefore, the prior art uses beam splitters as shown in FIGS. 27 and 28. Accordingly, when a non-polarized laser is used as a light source, together with a non-polarization type beam splitter, the amount of light is reduced to ¼ every time the laser beam travels via the wavefront converting element.
More specifically, the amount of light is reduced to ¼ in the process of illumination and also reduced to ¼ in the process of detection. That is, the amount of light is reduced to 1/16 in total. If a linearly polarized laser is used as a light source, together with a polarization beam splitter and a quarter-wave plate, the loss of light in the process of illumination can be prevented. However, in observation of fluorescence in a non-polarized state, the amount of light is reduced to ½ in the process of (fluorescence) detection.
Further, even when a polarization beam splitter and a quarter-wave plate are used as stated above, it is not always possible to use a linearly polarized laser as a light source. If a non-polarized laser is used to observe fluorescence, the amount of light is reduced to ½ in the process of illumination and also reduced to ½ in the process of detection. That is, the amount of light is reduced to ¼ in total.